5-Hydroxymethylcytosine and Globin Gene Switching

5-羟甲基胞嘧啶和珠蛋白基因转换

• 批准号: 8340878
• 负责人: DONALD LAVELLE
• 金额: $ 39.88万
• 依托单位: UNIVERSITY OF ILLINOIS AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-08-15 至 2016-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8340878
• 关键词: Adult; Agonist; Bone Marrow; CD34 gene; Cell Culture Techniques; Cells; Chemicals; Chromosomes, Artificial, Yeast; Clinical Trials; DNA; DNA Methylation; DNA Methyltransferase Inhibitor; Data; Development; Dimerization; Dioxygenases; Effectiveness; Enzyme Inhibitor Drugs; Erythroid Cells; Erythropoiesis; Eukaryotic Cell; Family; Fetal Hemoglobin; Fetal Liver; Fostering; Future; Gene Expression; Genes; Globin; Goals; Hemoglobin concentration result; Human; Knock-in Mouse; Knowledge; Laboratories; Lead; Longevity; Mediating; Modeling; Modification; Molecular; Mus; Papio; Pathway interactions; Patients; Pattern; Pharmaceutical Preparations; Physiological; Play; Pregnancy; Principal Investigator; Quality of life; RNA Interference; Regulation; Role; Series; Severities; Sickle Cell Anemia; Staging; Switch Genes; Symptoms; System; Testing; Thalassemia; Time; Transgenic Mice; Umbilical Cord Blood; Validation; beta Thalassemia; demethylation; design; erythroid differentiation; fetal; gamma Globin; mouse model; novel; programs; promoter; research study; therapy development

DESCRIPTION (provided by applicant): Increased fetal hemoglobin (HbF) levels lessen the severity of symptoms associated with sickle cell disease and beta-thalassemia and increase the quality of life and life span of patients. A complete understanding of the mechanism(s) responsible for developmental regulation of globin gene expression (globin gene switching) is important to develop new therapies to increase HbF. Over 30 years of experimental evidence from multiple laboratories strongly supports the hypothesis that DNA methylation plays a fundamental role in silencing expression of the gamma-globin gene in the adults. The use of pharmacological inhibitors of DNA methyltransferase to increase HbF levels was pioneered in the baboon model in our laboratory and led to a series of clinical trials that demonstrated the effectiveness of these drugs in patients. Using the baboon model, our laboratory also made the novel and important observation showing that the gamma-globin gene promoter was demethylated in a progressive manner during fetal liver erythropoiesis. The mechanism responsible for DNA demethylation of the gamma-globin promoter during fetal liver erythropoiesis remains unknown. Our preliminary data shows that increased levels of 5-hydroxymethylcytosine (5-hmC), a novel modification of 5-methylcytosine (5-mC) recently found in eukaryotic cells that is catalyzed by the TET dioxygenase family, are associated with the gamma-globin promoter in erythroid cells expressing elevated levels of gamma-globin. 5-hmC has been proposed to be a key intermediate in both passive and active mechanisms of DNA demethylation and therefore our preliminary data strongly suggests that 5-hmC is involved in the mechanism responsible for demethylation of the gamma-globin gene during fetal liver erythropoiesis. This proposal will investigate the hypothesis that DNA demethylation of the gamma-globin gene during fetal erythroid differentiation is accomplished through a TET- mediated pathway involving 5-hmC. Validation of this hypothesis will define a crucial mechanism (s) in normal developmental fetal stage-specific activation of gamma-globin expression. We envision that detailed knowledge of the normal, physiological mechanism responsible for gamma-globin gene demethylation during fetal liver erythropoiesis will foster the development of new strategies targeting this mechanism to achieve DNA demethylation and high level activation of the gamma-globin gene to increase HbF in patients. PUBLIC HEALTH RELEVANCE: Elevated fetal hemoglobin levels lessen the severity of sickle cell disease and beta-thalassemia and increase the lifespan of patients. A complete description of the mechanism responsible for the regulation of gamma-globin gene expression would be important in the design of future pharmacological and molecular therapies to increase HbF. DNA methylation is a key factor in the regulation of gamma-globin gene expression. The experiments described in this proposal seek to understand the role of 5-hydroxymethylcytosine in the mechanism(s) responsible for DNA demethylation of the gamma-globin promoter during fetal liver erythropoiesis. It is our long term goal to understand the mechanism responsible for developmental regulation of the globin gene expression as a means to developing therapies for sickle cell disease and beta-thalassemia. We expect that our studies will foster the development of new strategies targeting the normal, physiological mechanism responsible for gamma-globin gene DNA demethylation to achieve increased fetal hemoglobin levels in patients.

描述（由申请人提供）：增加的胎儿血红蛋白（HBF）水平减轻了与镰状细胞疾病和β-丘脑贫血相关的症状的严重程度，并提高了患者的生活质量和寿命。对负责球蛋白基因表达（Globin基因转换）发育调控的机制的完全理解对于开发新疗法以增加HBF很重要。来自多个实验室的30年来，有30年的实验证据强烈支持以下假设：DNA甲基化在沉默γ-球蛋白基因在成年人中的表达中起着至关重要的作用。在我们的实验室的狒狒模型中，使用了DNA甲基转移酶的药理抑制剂来升高HBF水平，并导致了一系列临床试验，这些试验证明了这些药物在患者中的有效性。使用狒狒模型，我们的实验室还做出了新颖而重要的观察结果，表明在胎儿肝红细胞生成期间以渐进式的方式将γ-球蛋白基因启动子脱甲基。胎儿肝脏红细胞生成期间γ-球蛋白启动子DNA脱甲基化的机制仍然未知。 Our preliminary data shows that increased levels of 5-hydroxymethylcytosine (5-hmC), a novel modification of 5-methylcytosine (5-mC) recently found in eukaryotic cells that is catalyzed by the TET dioxygenase family, are associated with the gamma-globin promoter in erythroid cells expressing elevated levels of gamma-globin.已经提出5-HMC是DNA脱甲基化的被动和主动机制中的关键中间体，因此我们的初步数据强烈表明5-HMC参与了负责在胎儿肝硬脂腺过程中脱甲基化的机制。该建议将研究以下假设：胎儿红细胞分化过程中γ-球蛋白基因的DNA脱甲基化通过涉及5-HMC的TET介导的途径来实现。该假设的验证将定义在正常发育性胎儿阶段特异性激活中的关键机制。我们设想，在胎儿肝脏红细胞生成过程中导致γ-球蛋白基因去甲基化的正常生理机制的详细了解将促进针对这种机制的新策略的发展，以实现DNA脱甲基化并实现患者HBF的HBF增加HBF的高水平脱甲基化。 公共卫生相关性：胎儿血红蛋白水平升高减轻了镰状细胞疾病和β-杂无血症的严重程度，并增加了患者的寿命。对负责调节γ-球蛋白基因表达的机制的完整描述对于增加HBF的未来药理和分子疗法的设计将很重要。 DNA甲基化是调节γ-球蛋白基因表达的关键因素。该提案中描述的实验旨在了解5-羟基甲基胞嘧啶在负责在胎儿肝红细胞生成过程中γ-珠蛋白启动子DNA脱甲基的机制中的作用。我们的长期目标是了解负责球蛋白基因表达的发育调节的机制，以此作为开发镰状细胞疾病和β-核阿无血症疗法的一种手段。我们预计，我们的研究将促进针对正常生理机制的新策略的发展，导致γ-球蛋白基因DNA脱甲基化，从而实现患者胎儿血红蛋白水平的提高。